CTRP5 over-expression in RPE cells leads to loss of RPE cell adhesion and retinal degeneration in wild-type mice

Abstract

Purpose: The complement C1q tumor necrosis factor-related protein-5 (C1QTNF5/CTRP5) is highly expressed in the RPE and cilliary epithelial layers in the eye. A pathogenic mutation, S163R, causes autosomal dominant late-onset retinal degeneration, with thick extracellular sub-RPE deposits, and choroidal neovascularization at later stages. Here, our goal was to test the effects of wild-type CTRP5 protein overexpression in RPE cells on retinal structure and function in normal mice.

Methods: We generated AAV vectors as tools to overexpress either CTRP5 or its dicistronic partner, the membrane-type frizzled related protein (MFRP), specifically in RPE cells of C57BL/6 mice. One microliter (10⁹ total vector genomes) of scAAV8 (Y733F) vectors containing an RPE-specific BEST1 promoter driving either a mouse Ctrp5 or Mfrp cDNA was injected subretinally into one eye of adult C57BL/6 mice, while the contralateral eyes were untreated and served as controls. Retinal function was assessed by full-field electroretinography (ERG) under scotopic (dark-adapted) conditions at 2 months post-injection. Pathological changes were investigated by optical coherence tomography, digital fundus imaging, evaluation of retinal morphology and immunohistochemistry.

Results: RPE targeted overexpression of CTRP5 leads to severe retinal degeneration, with pathological effects on both photoreceptors and RPE. The scAAV8 (Y733F)-CTRP5 treated eyes displayed a dramatic loss of photoreceptors over the entire retina. In addition, the RPE cells appeared to have lost their intercellular junctional integrity, and separated from their lateral neighbor in the RPE monolayer. Some areas of CTRP5 treatment suffered complete loss of both RPE and photoreceptor cells by 3 months post-injection. In contrast, scAAV8 (Y733F)-MFRP treatment led to no detrimental effects on retinal structure or function, with the RPE appearing healthy even with robust overexpression of MFRP within the apical membrane and its microvilli.

Conclusions: Abnormally increased levels of CTRP5 in RPE lead to widespread photoreceptor cell death, and loss of RPE intercellular adhesion. This suggests that CTRP5 may play a key role in modulating the RPE intercellular tight junctions, and its excess may contribute to the progression of choroidal neovascularization (CNV) in pathological conditions such as age related macular degeneration (AMD).

Keywords: 412 age-related macular degeneration • 648 photoreceptors • 446 cell adhesions/cell junctions